Rib-X Pharmaceuticals, Inc. has proprietary, high-resolution structural information on how antibiotics bind to the large ribosomal subunit (50S). Using this information in combination with a computational approach that calculates the energetics of RNA-ligand interactions and drug-like properties of these antibiotics, a series of novel compounds were designed and synthesized. One of the compounds was complexed to Haloarcula marismortui 50S ribosomal subunits and solved to 3.0 Angstroms resolution. The position of the analog within the 50S subunit validated our approach and suggested that solving related structures within the series would provide a solid platform to understand how this series of compounds overcomes target-based resistance. To aid in drug design, this proposal seeks to: 1. Determine two additional high resolution structures of compounds complexed with the 50S subunit of Haloarcula marismortui. 2. Determine the minimally-active chemical core in the compound series. 3. Determine the pharmacokinetics of the most promising compound in mice. The overall goal of this work is to identify new chemical entities for the treatment of antibiotic resistant bacterial infections. This is important because the incidence of antimicrobial resistance has increased in hospital and community settings. Therapeutic failures with current agents, extended hospital stays, and the use of increasingly costly and toxic antimicrobials that contribute to healthcare costs is driving the need for new antimicrobial agents that overcome multidrug resistance. Resistance is likely to become worse when patents for widely used, branded antibiotics like Augmentin, Cipro, and Zithromax expire, resulting in widespread use of generic versions of these drugs. The 50S subunit is a highly validated target in that two of the top four oral antibiotics in the US target it (Zithromax and Biaxen); collectively, these two antibiotics account for >$2 billion in annual sales. Further, the most recently approved class of antibiotics, Zyvox, also binds to the 50S. Zyvox is used to treat serious Gram-positive nosocomial infections. Thus, the 50S presents an opportunity to target medical needs and to find new antimicrobials for both community- and hospital-acquired infections.